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Dos Santos Bronel BA, Maquigussa E, Boim MA, da Silva Novaes A. Effect of extracellular vesicles derived from induced pluripotent stem cells on mesangial cells underwent a model of fibrosis in vitro. Sci Rep 2023; 13:15749. [PMID: 37735602 PMCID: PMC10514265 DOI: 10.1038/s41598-023-42912-6] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/15/2023] [Accepted: 09/15/2023] [Indexed: 09/23/2023] Open
Abstract
The fibrogenic process plays a significant pathophysiological role in the progression of chronic kidney disease. Inhibition of the renin-angiotensin system (RAS) is one strategy to delay disease progression but does not reverse established fibrosis. In this context, induced pluripotent stem cells (iPSCs) have been considered an alternative due to their regenerative potential. iPSCs exert their effects through paracrine signaling, which releases specific biomolecules into the extracellular environment, either directly or within extracellular vesicle (EVs), that can reach target cells. This study aims to evaluate the potential beneficial effects of iPSC-derived EVs (EV-iPSCs) in an in vitro model of fibrosis using mouse mesangial cells (MMCs) stimulated with TGF-β. EV-iPSCs were obtained by differentially ultracentrifuging iPSCs culture medium. MMCs were stimulated with 5 ng/mL of TGF-β and simultaneously treated with or without EV-iPSCs for 24 h. Markers of inflammation, fibrosis, and RAS components were assessed using RT-PCR, western blotting, and immunofluorescence. Under TGF-β stimulus, MMCs exhibited increased expression of inflammation markers, RAS components, and fibrosis. However, these changes were mitigated in the presence of EV-iPSCs. EV-iPSCs effectively reduced inflammation, RAS activation, and fibrogenesis in this fibrosis model involving mesangial cells, suggesting their potential as a strategy to reduce glomerular sclerosis.
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Affiliation(s)
- Bruno Aristides Dos Santos Bronel
- Renal Division, Department of Medicine, Universidade Federal de São Paulo, 781 Pedro de Toledo St, 13° Floor, São Paulo, SP, 04039-032, Brazil
| | - Edgar Maquigussa
- Renal Division, Department of Medicine, Universidade Federal de São Paulo, 781 Pedro de Toledo St, 13° Floor, São Paulo, SP, 04039-032, Brazil
| | - Mirian Aparecida Boim
- Renal Division, Department of Medicine, Universidade Federal de São Paulo, 781 Pedro de Toledo St, 13° Floor, São Paulo, SP, 04039-032, Brazil
| | - Antônio da Silva Novaes
- Renal Division, Department of Medicine, Universidade Federal de São Paulo, 781 Pedro de Toledo St, 13° Floor, São Paulo, SP, 04039-032, Brazil.
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Matos BMD, Stimamiglio MA, Correa A, Robert AW. Human pluripotent stem cell-derived extracellular vesicles: From now to the future. World J Stem Cells 2023; 15:453-465. [PMID: 37342215 PMCID: PMC10277970 DOI: 10.4252/wjsc.v15.i5.453] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/26/2022] [Revised: 02/14/2023] [Accepted: 04/13/2023] [Indexed: 05/26/2023] Open
Abstract
Extracellular vesicles (EVs) are nanometric particles that enclose cell-derived bioactive molecules in a lipid bilayer and serve as intercellular communication tools. Accordingly, in various biological contexts, EVs are reported to engage in immune modulation, senescence, and cell proliferation and differentiation. Therefore, EVs could be key elements for potential off-the-shelf cell-free therapy. Little has been studied regarding EVs derived from human pluripotent stem cells (hPSC-EVs), even though hPSCs offer good opportunities for induction of tissue regeneration and unlimited proliferative ability. In this review article, we provide an overview of studies using hPSC-EVs, focusing on identifying the conditions in which the cells are cultivated for the isolation of EVs, how they are characterized, and applications already demonstrated. The topics reported in this article highlight the incipient status of the studies in the field and the significance of hPSC-EVs’ prospective applications as PSC-derived cell-free therapy products.
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Affiliation(s)
- Bruno Moises de Matos
- Stem Cells Basic Biology Laboratory, Carlos Chagas Institute, Curitiba 81350010, Paraná, Brazil
| | | | - Alejandro Correa
- Stem Cells Basic Biology Laboratory, Carlos Chagas Institute, Curitiba 81350010, Paraná, Brazil
| | - Anny Waloski Robert
- Stem Cells Basic Biology Laboratory, Carlos Chagas Institute, Curitiba 81350010, Paraná, Brazil
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3
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Louro AF, Paiva MA, Oliveira MR, Kasper KA, Alves PM, Gomes‐Alves P, Serra M. Bioactivity and miRNome Profiling of Native Extracellular Vesicles in Human Induced Pluripotent Stem Cell-Cardiomyocyte Differentiation. ADVANCED SCIENCE (WEINHEIM, BADEN-WURTTEMBERG, GERMANY) 2022; 9:e2104296. [PMID: 35322574 PMCID: PMC9130911 DOI: 10.1002/advs.202104296] [Citation(s) in RCA: 13] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/27/2021] [Revised: 01/05/2022] [Indexed: 05/17/2023]
Abstract
Extracellular vesicles (EV) are an attractive therapy to boost cardiac regeneration. Nevertheless, identification of native EV and corresponding cell platform(s) suitable for therapeutic application, is still a challenge. Here, EV are isolated from key stages of the human induced pluripotent stem cell-cardiomyocyte (hiPSC-CM) differentiation and maturation, i.e., from hiPSC (hiPSC-EV), cardiac progenitors, immature and mature cardiomyocytes, with the aim of identifying a promising cell biofactory for EV production, and pinpoint the genetic signatures of bioactive EV. EV secreted by hiPSC and cardiac derivatives show a typical size distribution profile and the expression of specific EV markers. Bioactivity assays show increased tube formation and migration in HUVEC treated with hiPSC-EV compared to EV from committed cell populations. hiPSC-EV also significantly increase cell cycle activity of hiPSC-CM. Global miRNA expression profiles, obtained by small RNA-seq analysis, corroborate an EV-miRNA pattern indicative of stem cell to cardiomyocyte specification, confirming that hiPSC-EV are enriched in pluripotency-associated miRNA with higher in vitro pro-angiogenic and pro-proliferative properties. In particular, a stemness maintenance miRNA cluster upregulated in hiPSC-EV targets the PTEN/PI3K/AKT pathway, involved in cell proliferation and survival. Overall, the findings validate hiPSC as cell biofactories for EV production for cardiac regenerative applications.
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Affiliation(s)
- Ana F. Louro
- iBETInstituto de Biologia Experimental e TecnológicaApartado 12Oeiras2781‐901Portugal
- ITQB‐NOVAInstituto de Tecnologia Química e Biológica António XavierUniversidade Nova de LisboaAv. da RepúblicaOeiras2780‐157Portugal
| | - Marta A. Paiva
- iBETInstituto de Biologia Experimental e TecnológicaApartado 12Oeiras2781‐901Portugal
- ITQB‐NOVAInstituto de Tecnologia Química e Biológica António XavierUniversidade Nova de LisboaAv. da RepúblicaOeiras2780‐157Portugal
| | - Marta R. Oliveira
- iBETInstituto de Biologia Experimental e TecnológicaApartado 12Oeiras2781‐901Portugal
- ITQB‐NOVAInstituto de Tecnologia Química e Biológica António XavierUniversidade Nova de LisboaAv. da RepúblicaOeiras2780‐157Portugal
| | - Katharina A. Kasper
- iBETInstituto de Biologia Experimental e TecnológicaApartado 12Oeiras2781‐901Portugal
- ITQB‐NOVAInstituto de Tecnologia Química e Biológica António XavierUniversidade Nova de LisboaAv. da RepúblicaOeiras2780‐157Portugal
| | - Paula M. Alves
- iBETInstituto de Biologia Experimental e TecnológicaApartado 12Oeiras2781‐901Portugal
- ITQB‐NOVAInstituto de Tecnologia Química e Biológica António XavierUniversidade Nova de LisboaAv. da RepúblicaOeiras2780‐157Portugal
| | - Patrícia Gomes‐Alves
- iBETInstituto de Biologia Experimental e TecnológicaApartado 12Oeiras2781‐901Portugal
- ITQB‐NOVAInstituto de Tecnologia Química e Biológica António XavierUniversidade Nova de LisboaAv. da RepúblicaOeiras2780‐157Portugal
| | - Margarida Serra
- iBETInstituto de Biologia Experimental e TecnológicaApartado 12Oeiras2781‐901Portugal
- ITQB‐NOVAInstituto de Tecnologia Química e Biológica António XavierUniversidade Nova de LisboaAv. da RepúblicaOeiras2780‐157Portugal
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Tracy EP, Stielberg V, Rowe G, Benson D, Nunes SS, Hoying JB, Murfee WL, LeBlanc AJ. State of the field: cellular and exosomal therapeutic approaches in vascular regeneration. Am J Physiol Heart Circ Physiol 2022; 322:H647-H680. [PMID: 35179976 PMCID: PMC8957327 DOI: 10.1152/ajpheart.00674.2021] [Citation(s) in RCA: 15] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/14/2021] [Revised: 02/07/2022] [Accepted: 02/09/2022] [Indexed: 01/19/2023]
Abstract
Pathologies of the vasculature including the microvasculature are often complex in nature, leading to loss of physiological homeostatic regulation of patency and adequate perfusion to match tissue metabolic demands. Microvascular dysfunction is a key underlying element in the majority of pathologies of failing organs and tissues. Contributing pathological factors to this dysfunction include oxidative stress, mitochondrial dysfunction, endoplasmic reticular (ER) stress, endothelial dysfunction, loss of angiogenic potential and vascular density, and greater senescence and apoptosis. In many clinical settings, current pharmacologic strategies use a single or narrow targeted approach to address symptoms of pathology rather than a comprehensive and multifaceted approach to address their root cause. To address this, efforts have been heavily focused on cellular therapies and cell-free therapies (e.g., exosomes) that can tackle the multifaceted etiology of vascular and microvascular dysfunction. In this review, we discuss 1) the state of the field in terms of common therapeutic cell population isolation techniques, their unique characteristics, and their advantages and disadvantages, 2) common molecular mechanisms of cell therapies to restore vascularization and/or vascular function, 3) arguments for and against allogeneic versus autologous applications of cell therapies, 4) emerging strategies to optimize and enhance cell therapies through priming and preconditioning, and, finally, 5) emerging strategies to bolster therapeutic effect. Relevant and recent clinical and animal studies using cellular therapies to restore vascular function or pathologic tissue health by way of improved vascularization are highlighted throughout these sections.
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Affiliation(s)
- Evan Paul Tracy
- Cardiovascular Innovation Institute and the Department of Physiology, University of Louisville, Louisville, Kentucky
| | - Virginia Stielberg
- Cardiovascular Innovation Institute and the Department of Physiology, University of Louisville, Louisville, Kentucky
| | - Gabrielle Rowe
- Cardiovascular Innovation Institute and the Department of Physiology, University of Louisville, Louisville, Kentucky
| | - Daniel Benson
- Cardiovascular Innovation Institute and the Department of Physiology, University of Louisville, Louisville, Kentucky
- Department of Bioengineering, University of Louisville, Louisville, Kentucky
| | - Sara S Nunes
- Toronto General Hospital Research Institute, University Health Network, Toronto, Ontario, Canada
- Institute of Biomedical Engineering, University of Toronto, Toronto, Ontario, Canada
- Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada
- Heart & Stroke/Richard Lewar Centre of Excellence, University of Toronto, Toronto, Ontario, Canada
| | - James B Hoying
- Advanced Solutions Life Sciences, Manchester, New Hampshire
| | - Walter Lee Murfee
- J. Crayton Pruitt Family Department of Biomedical Engineering, University of Florida, Gainesville, Florida
| | - Amanda Jo LeBlanc
- Cardiovascular Innovation Institute and the Department of Physiology, University of Louisville, Louisville, Kentucky
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Andrade AC, Wolf M, Binder HM, Gomes FG, Manstein F, Ebner-Peking P, Poupardin R, Zweigerdt R, Schallmoser K, Strunk D. Hypoxic Conditions Promote the Angiogenic Potential of Human Induced Pluripotent Stem Cell-Derived Extracellular Vesicles. Int J Mol Sci 2021; 22:ijms22083890. [PMID: 33918735 PMCID: PMC8070165 DOI: 10.3390/ijms22083890] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/26/2021] [Revised: 04/06/2021] [Accepted: 04/07/2021] [Indexed: 12/11/2022] Open
Abstract
Stem cells secrete paracrine factors including extracellular vesicles (EVs) which can mediate cellular communication and support the regeneration of injured tissues. Reduced oxygen (hypoxia) as a key regulator in development and regeneration may influence cellular communication via EVs. We asked whether hypoxic conditioning during human induced pluripotent stem cell (iPSC) culture effects their EV quantity, quality or EV-based angiogenic potential. We produced iPSC-EVs from large-scale culture-conditioned media at 1%, 5% and 18% air oxygen using tangential flow filtration (TFF), with or without subsequent concentration by ultracentrifugation (TUCF). EVs were quantified by tunable resistive pulse sensing (TRPS), characterized according to MISEV2018 guidelines, and analyzed for angiogenic potential. We observed superior EV recovery by TFF compared to TUCF. We confirmed hypoxia efficacy by HIF-1α stabilization and pimonidazole hypoxyprobe. EV quantity did not differ significantly at different oxygen conditions. Significantly elevated angiogenic potential was observed for iPSC-EVs derived from 1% oxygen culture by TFF or TUCF as compared to EVs obtained at higher oxygen or the corresponding EV-depleted soluble factor fractions. Data thus demonstrate that cell-culture oxygen conditions and mode of EV preparation affect iPSC-EV function. We conclude that selecting appropriate protocols will further improve production of particularly potent iPSC-EV-based therapeutics.
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Affiliation(s)
- André Cronemberger Andrade
- Cell Therapy Institute, Spinal Cord Injury and Tissue Regeneration Center Salzburg (SCI-TReCS), Paracelsus Medical University (PMU), 5020 Salzburg, Austria; (A.C.A.); (M.W.); (H.-M.B.); (P.E.-P.); (R.P.)
| | - Martin Wolf
- Cell Therapy Institute, Spinal Cord Injury and Tissue Regeneration Center Salzburg (SCI-TReCS), Paracelsus Medical University (PMU), 5020 Salzburg, Austria; (A.C.A.); (M.W.); (H.-M.B.); (P.E.-P.); (R.P.)
| | - Heide-Marie Binder
- Cell Therapy Institute, Spinal Cord Injury and Tissue Regeneration Center Salzburg (SCI-TReCS), Paracelsus Medical University (PMU), 5020 Salzburg, Austria; (A.C.A.); (M.W.); (H.-M.B.); (P.E.-P.); (R.P.)
| | - Fausto Gueths Gomes
- Department of Transfusion Medicine and SCI-TReCS, Paracelsus Medical University (PMU), 5020 Salzburg, Austria; (F.G.G.); (K.S.)
| | - Felix Manstein
- Department of Cardiac, Thoracic, Transplantation and Vascular Surgery, Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Hannover Medical School, 30625 Hannover, Germany; (F.M.); (R.Z.)
| | - Patricia Ebner-Peking
- Cell Therapy Institute, Spinal Cord Injury and Tissue Regeneration Center Salzburg (SCI-TReCS), Paracelsus Medical University (PMU), 5020 Salzburg, Austria; (A.C.A.); (M.W.); (H.-M.B.); (P.E.-P.); (R.P.)
| | - Rodolphe Poupardin
- Cell Therapy Institute, Spinal Cord Injury and Tissue Regeneration Center Salzburg (SCI-TReCS), Paracelsus Medical University (PMU), 5020 Salzburg, Austria; (A.C.A.); (M.W.); (H.-M.B.); (P.E.-P.); (R.P.)
| | - Robert Zweigerdt
- Department of Cardiac, Thoracic, Transplantation and Vascular Surgery, Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Hannover Medical School, 30625 Hannover, Germany; (F.M.); (R.Z.)
| | - Katharina Schallmoser
- Department of Transfusion Medicine and SCI-TReCS, Paracelsus Medical University (PMU), 5020 Salzburg, Austria; (F.G.G.); (K.S.)
| | - Dirk Strunk
- Cell Therapy Institute, Spinal Cord Injury and Tissue Regeneration Center Salzburg (SCI-TReCS), Paracelsus Medical University (PMU), 5020 Salzburg, Austria; (A.C.A.); (M.W.); (H.-M.B.); (P.E.-P.); (R.P.)
- Correspondence:
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6
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Wang AYL. Human Induced Pluripotent Stem Cell-Derived Exosomes as a New Therapeutic Strategy for Various Diseases. Int J Mol Sci 2021; 22:1769. [PMID: 33578948 PMCID: PMC7916646 DOI: 10.3390/ijms22041769] [Citation(s) in RCA: 29] [Impact Index Per Article: 7.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/31/2020] [Revised: 02/01/2021] [Accepted: 02/07/2021] [Indexed: 02/07/2023] Open
Abstract
Recently, an increasing number of studies have demonstrated that induced pluripotent stem cells (iPSCs) and iPSC-derived cells display therapeutic effects, mainly via the paracrine mechanism in addition to their transdifferentiation ability. Exosomes have emerged as an important paracrine factor for iPSCs to repair injured cells through the delivery of bioactive components. Animal reports of iPSC-derived exosomes on various disease models are increasing, such as in heart, limb, liver, skin, bone, eye and neurological disease and so forth. This review aims to summarize the therapeutic effects of iPSC-derived exosomes on various disease models and their properties, such as angiogenesis, cell proliferation and anti-apoptosis, with the hopes of improving their potential role in clinical applications and functional restoration.
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Affiliation(s)
- Aline Yen Ling Wang
- Center for Vascularized Composite Allotransplantation, Chang Gung Memorial Hospital, 5 Fu-hsing Street, Gueishan, Taoyuan 333, Taiwan
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7
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Marzano M, Bou-Dargham MJ, Cone AS, York S, Helsper S, Grant SC, Meckes DG, Sang QXA, Li Y. Biogenesis of Extracellular Vesicles Produced from Human-Stem-Cell-Derived Cortical Spheroids Exposed to Iron Oxides. ACS Biomater Sci Eng 2021; 7:1111-1122. [PMID: 33525864 DOI: 10.1021/acsbiomaterials.0c01286] [Citation(s) in RCA: 18] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/28/2022]
Abstract
Stem-cell-derived extracellular vesicles (EVs) are promising tools for therapeutic delivery and imaging in the medical research fields. EVs that arise from endosomal compartments or plasma membrane budding consist of exosomes and microvesicles, which range between 30 and 200 nm and 100-1000 nm, respectively. Iron oxide nanoparticles can be used to label stem cells or possibly EVs for magnetic resonance imaging. This could be a novel way to visualize areas in the body that are affected by neurological disorders such as stroke. Human induced pluripotent stem cells (iPSK3 cells) were plated on low-attachment plates and treated with SB431542 and LDN193189 during the first week for the induction of cortical spheroid formation and grown with fibroblast growth factor 2 and cyclopamine during the second week for the neural progenitor cell (iNPC) differentiation. iNPCs were then grown on attachment plates and treated with iron oxide (Fe3O4) nanoparticles at different sizes (8, 15, and 30 nm in diameter) and concentrations (0.1, 10, and 100 μM). The spheroids and media collected from these cultures were used for iron oxide detection as well as EV isolation and characterizations, respectively. MTT assay demonstrated that the increased size and concentration of the iron oxide nanoparticles had little effect on the metabolic activity of iNPCs. In addition, the Live/Dead assay showed high viability in all the nanoparticle treated groups and the untreated control. The EVs isolated from these culture groups were analyzed and displayed similar or higher EV counts compared with control. The observed EV size averaged 200-250 nm, and electron microscopy revealed the expected exosome morphology for EVs from all groups. RT-PCR analysis of EV biogenesis markers (CD63, CD81, Alix, TSG101, Syntenin1, ADAM10, RAB27b, and Syndecan) showed differential expression between the iron-oxide-treated cultures and nontreated cultures, as well as between adherent and nonadherent 3D cultures. Iron oxide nanoparticles were detected inside the cortical spheroid cells but not EVs by MRI. The addition of iron oxide nanoparticles does not induce significant cytotoxic effects to cortical spheroids. In addition,, nanoparticles may stimulate the biogenesis of EVs when added to cortical spheroids in vitro.
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Affiliation(s)
- Mark Marzano
- Department of Chemical and Biomedical Engineering, Florida State University, Tallahassee, Florida 32306, United States
| | - Mayassa J Bou-Dargham
- Department of Chemistry and Biochemistry, Florida State University, Tallahassee, Florida 32306, United States
| | - Allaura S Cone
- Department of Biomedical Sciences, College of Medicine, Florida State University, Tallahassee, Florida 32304, United States
| | - Sara York
- Department of Biomedical Sciences, College of Medicine, Florida State University, Tallahassee, Florida 32304, United States
| | - Shannon Helsper
- Department of Chemical and Biomedical Engineering, Florida State University, Tallahassee, Florida 32306, United States.,The National High Magnetic Field Laboratory, Florida State University, Tallahassee, Florida 32310, United States
| | - Samuel C Grant
- Department of Chemical and Biomedical Engineering, Florida State University, Tallahassee, Florida 32306, United States.,The National High Magnetic Field Laboratory, Florida State University, Tallahassee, Florida 32310, United States
| | - David G Meckes
- Department of Biomedical Sciences, College of Medicine, Florida State University, Tallahassee, Florida 32304, United States
| | - Qing-Xiang Amy Sang
- Department of Chemistry and Biochemistry, Florida State University, Tallahassee, Florida 32306, United States
| | - Yan Li
- Department of Chemical and Biomedical Engineering, Florida State University, Tallahassee, Florida 32306, United States
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Ha DH, Kim HK, Lee J, Kwon HH, Park GH, Yang SH, Jung JY, Choi H, Lee JH, Sung S, Yi YW, Cho BS. Mesenchymal Stem/Stromal Cell-Derived Exosomes for Immunomodulatory Therapeutics and Skin Regeneration. Cells 2020; 9:E1157. [PMID: 32392899 PMCID: PMC7290908 DOI: 10.3390/cells9051157] [Citation(s) in RCA: 304] [Impact Index Per Article: 60.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/20/2020] [Revised: 04/25/2020] [Accepted: 05/04/2020] [Indexed: 12/12/2022] Open
Abstract
Exosomes are nano-sized vesicles that serve as mediators for cell-to-cell communication. With their unique nucleic acids, proteins, and lipids cargo compositions that reflect the characteristics of producer cells, exosomes can be utilized as cell-free therapeutics. Among exosomes derived from various cellular origins, mesenchymal stem cell-derived exosomes (MSC-exosomes) have gained great attention due to their immunomodulatory and regenerative functions. Indeed, many studies have shown anti-inflammatory, anti-aging and wound healing effects of MSC-exosomes in various in vitro and in vivo models. In addition, recent advances in the field of exosome biology have enabled development of specific guidelines and quality control methods, which will ultimately lead to clinical application of exosomes. This review highlights recent studies that investigate therapeutic potential of MSC-exosomes and relevant mode of actions for skin diseases, as well as quality control measures required for development of exosome-derived therapeutics.
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Affiliation(s)
- Dae Hyun Ha
- ExoCoBio Exosome Institute (EEI), ExoCoBio Inc., Seoul 08594, Korea; (D.H.H.); (H.-k.K.); (J.H.L.); (S.S.)
| | - Hyun-keun Kim
- ExoCoBio Exosome Institute (EEI), ExoCoBio Inc., Seoul 08594, Korea; (D.H.H.); (H.-k.K.); (J.H.L.); (S.S.)
| | - Joon Lee
- School of Chemical and Biological Engineering, Seoul National University, Seoul 08826, Korea;
| | | | - Gyeong-Hun Park
- Department of Dermatology, Dongtan Sacred Heart Hospital, Hallym University College of Medicine, Hwasweong-si, Gyeonggi-do 18450, Korea;
| | | | | | | | - Jun Ho Lee
- ExoCoBio Exosome Institute (EEI), ExoCoBio Inc., Seoul 08594, Korea; (D.H.H.); (H.-k.K.); (J.H.L.); (S.S.)
| | - Sumi Sung
- ExoCoBio Exosome Institute (EEI), ExoCoBio Inc., Seoul 08594, Korea; (D.H.H.); (H.-k.K.); (J.H.L.); (S.S.)
| | - Yong Weon Yi
- ExoCoBio Exosome Institute (EEI), ExoCoBio Inc., Seoul 08594, Korea; (D.H.H.); (H.-k.K.); (J.H.L.); (S.S.)
| | - Byong Seung Cho
- ExoCoBio Exosome Institute (EEI), ExoCoBio Inc., Seoul 08594, Korea; (D.H.H.); (H.-k.K.); (J.H.L.); (S.S.)
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9
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Jeske R, Bejoy J, Marzano M, Li Y. Human Pluripotent Stem Cell-Derived Extracellular Vesicles: Characteristics and Applications. TISSUE ENGINEERING. PART B, REVIEWS 2020; 26:129-144. [PMID: 31847715 PMCID: PMC7187972 DOI: 10.1089/ten.teb.2019.0252] [Citation(s) in RCA: 33] [Impact Index Per Article: 6.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/15/2019] [Accepted: 12/16/2019] [Indexed: 02/06/2023]
Abstract
Extracellular vesicles (EVs), including exosomes and microvesicles, are found to play an important role in various biological processes and maintaining tissue homeostasis. Because of the protective effects, stem cell-derived EVs can be used to reduce oxidative stress and apoptosis in the recipient cells. In addition, EVs/exosomes have been used as directional communication tools between stem cells and parenchymal cells, giving them the ability to serve as biomarkers. Likewise, altered EVs/exosomes can be utilized for drug delivery by loading with proteins, small interfering RNAs, and viral vectors, in particular, because EVs/exosomes are able to cross the blood-brain barrier. In this review article, the properties of human induced pluripotent stem cell (iPSC)-derived EVs are discussed. The biogenesis, that is, how EVs originate in the endosomal compartment or from the cell layer of microvesicles, EV composition, the available methods of purification, and characterizations of EVs/exosomes are summarized. In particular, EVs/exosomes derived from iPSCs of different lineage specifications and the applications of these stem cell-derived exosomes in neurological diseases are discussed. Impact statement In this review, we summarized the work related to extracellular vesicles (EVs) derived from human pluripotent stem cells (hPSCs). In particular, EVs/exosomes derived from hPSCs of different lineage specifications and the applications of these stem cell-derived exosomes in neurological diseases are discussed. The results highlight the important role of cell-cell interactions in neural cellular phenotype and neurodegeneration. The findings reported in this article are significant for pluripotent stem cell-derived cell-free products toward applications in stem cell-based therapies.
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Affiliation(s)
- Richard Jeske
- Department of Chemical and Biomedical Engineering, FAMU-FSU College of Engineering, Florida State University, Tallahassee, Florida
| | - Julie Bejoy
- Department of Chemical and Biomedical Engineering, FAMU-FSU College of Engineering, Florida State University, Tallahassee, Florida
| | - Mark Marzano
- Department of Chemical and Biomedical Engineering, FAMU-FSU College of Engineering, Florida State University, Tallahassee, Florida
| | - Yan Li
- Department of Chemical and Biomedical Engineering, FAMU-FSU College of Engineering, Florida State University, Tallahassee, Florida
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10
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Fan C, Zhang E, Joshi J, Yang J, Zhang J, Zhu W. Utilization of Human Induced Pluripotent Stem Cells for Cardiac Repair. Front Cell Dev Biol 2020; 8:36. [PMID: 32117968 PMCID: PMC7025514 DOI: 10.3389/fcell.2020.00036] [Citation(s) in RCA: 29] [Impact Index Per Article: 5.8] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/30/2019] [Accepted: 01/15/2020] [Indexed: 12/18/2022] Open
Abstract
The paracrine effect, mediated by chemical signals that induce a physiological response on neighboring cells in the same tissue, is an important regenerative mechanism for stem cell-based therapy. Exosomes are cell-secreted nanovesicles (50-120 nm) of endosomal origin, and have been demonstrated to be a major contributor to the observed stem cell-mediated paracrine effect in the cardiac repair process. Following cardiac injury, exosomes deriving from exogenous stem cells have been shown to regulate cell apoptosis, proliferation, angiogenesis, and fibrosis in the infarcted heart. Exosomes also play a crucial role in the intercellular communication between donor and recipient cells. Human induced pluripotent stem cells (hiPSCs) are promising cell sources for autologous cell therapy in regenerative medicine. Here, we review recent advances in the field of progenitor-cell derived, exosome-based cardiac repair, with special emphasis on exosomes derived from hiPSCs.
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Affiliation(s)
- Chengming Fan
- Department of Biomedical Engineering, School of Medicine and School of Engineering, University of Alabama at Birmingham, Birmingham, AL, United States
- Department of Cardiovascular Surgery, The Second Xiangya Hospital, Central South University, Changsha, China
| | - Eric Zhang
- Department of Biomedical Engineering, School of Medicine and School of Engineering, University of Alabama at Birmingham, Birmingham, AL, United States
| | - Jyotsna Joshi
- Department of Cardiovascular Medicine, Physiology and Biomedical Engineering, Mayo Clinic, Scottsdale, AZ, United States
| | - Jinfu Yang
- Department of Cardiovascular Surgery, The Second Xiangya Hospital, Central South University, Changsha, China
| | - Jianyi Zhang
- Department of Biomedical Engineering, School of Medicine and School of Engineering, University of Alabama at Birmingham, Birmingham, AL, United States
| | - Wuqiang Zhu
- Department of Cardiovascular Medicine, Physiology and Biomedical Engineering, Mayo Clinic, Scottsdale, AZ, United States
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11
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Lee YXF, Johansson H, Wood MJA, El Andaloussi S. Considerations and Implications in the Purification of Extracellular Vesicles - A Cautionary Tale. Front Neurosci 2019; 13:1067. [PMID: 31680809 PMCID: PMC6813730 DOI: 10.3389/fnins.2019.01067] [Citation(s) in RCA: 38] [Impact Index Per Article: 6.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/08/2019] [Accepted: 09/24/2019] [Indexed: 12/29/2022] Open
Abstract
Extracellular vesicles (EVs) are nano-sized particles constitutively released from cells into all biological fluids. Interestingly, these vesicles contain genetic cargoes including proteins, RNA and bioactive lipids that can be functionally delivered and affect recipient cells. As a result, there is growing interest in studying EVs in pathological conditions, including central nervous system (CNS)-related diseases, as EVs may be used for diagnostic purposes or as therapeutic agents. However, one major bottleneck is the need for better EV purification strategies when considering complex biological sources such as serum/protein-rich media or plasma. In this study, we have performed a systematic comparison study between the current gold-standard method: ultracentrifugation, to an alternative: size-exclusion chromatography (LC), using induced pluripotent stem cell (iPSC) derived complex media as a model system. We demonstrate that LC allows for derivation of purer EVs from iPSCs, which was previously impossible with the original UC method. Importantly, our study further highlights the various drawbacks when using the conventional UC approach that lead to misinterpretation of EV data. Lastly, we describe novel data on our iPSC-EVs; how they could relate to stem cell biology and discuss their potential use as EV therapeutics for CNS diseases.
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Affiliation(s)
- Yi Xin Fiona Lee
- Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, United Kingdom.,Genome Institute of Singapore, Agency for Science, Technology and Research, Singapore, Singapore
| | - Henrik Johansson
- Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Karolinska Institutet, Stockholm, Sweden
| | - Matthew J A Wood
- Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, United Kingdom
| | - Samir El Andaloussi
- Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, United Kingdom.,Department of Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden
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12
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Differential Effects of Extracellular Vesicles of Lineage-Specific Human Pluripotent Stem Cells on the Cellular Behaviors of Isogenic Cortical Spheroids. Cells 2019; 8:cells8090993. [PMID: 31466320 PMCID: PMC6770916 DOI: 10.3390/cells8090993] [Citation(s) in RCA: 20] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/26/2019] [Revised: 08/23/2019] [Accepted: 08/27/2019] [Indexed: 02/07/2023] Open
Abstract
Extracellular vesicles (EVs) contribute to a variety of signaling processes and the overall physiological and pathological states of stem cells and tissues. Human induced pluripotent stem cells (hiPSCs) have unique characteristics that can mimic embryonic tissue development. There is growing interest in the use of EVs derived from hiPSCs as therapeutics, biomarkers, and drug delivery vehicles. However, little is known about the characteristics of EVs secreted by hiPSCs and paracrine signaling during tissue morphogenesis and lineage specification. Methods: In this study, the physical and biological properties of EVs isolated from hiPSC-derived neural progenitors (ectoderm), hiPSC-derived cardiac cells (mesoderm), and the undifferentiated hiPSCs (healthy iPSK3 and Alzheimer’s-associated SY-UBH lines) were analyzed. Results: Nanoparticle tracking analysis and electron microscopy results indicate that hiPSC-derived EVs have an average size of 100–250 nm. Immunoblot analyses confirmed the enrichment of exosomal markers Alix, CD63, TSG101, and Hsc70 in the purified EV preparations. MicroRNAs including miR-133, miR-155, miR-221, and miR-34a were differently expressed in the EVs isolated from distinct hiPSC lineages. Treatment of cortical spheroids with hiPSC-EVs in vitro resulted in enhanced cell proliferation (indicated by BrdU+ cells) and axonal growth (indicated by β-tubulin III staining). Furthermore, hiPSC-derived EVs exhibited neural protective abilities in Aβ42 oligomer-treated cultures, enhancing cell viability and reducing oxidative stress. Our results demonstrate that the paracrine signaling provided by tissue context-dependent EVs derived from hiPSCs elicit distinct responses to impact the physiological state of cortical spheroids. Overall, this study advances our understanding of cell‒cell communication in the stem cell microenvironment and provides possible therapeutic options for treating neural degeneration.
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13
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Branscome H, Paul S, Khatkar P, Kim Y, Barclay RA, Pinto DO, Yin D, Zhou W, Liotta LA, El-Hage N, Kashanchi F. Stem Cell Extracellular Vesicles and their Potential to Contribute to the Repair of Damaged CNS Cells. J Neuroimmune Pharmacol 2019; 15:520-537. [PMID: 31338754 DOI: 10.1007/s11481-019-09865-y] [Citation(s) in RCA: 26] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/17/2019] [Accepted: 07/10/2019] [Indexed: 12/31/2022]
Abstract
Neurological diseases and disorders are leading causes of death and disability worldwide. Many of these pathologies are associated with high levels of neuroinflammation and irreparable tissue damage. As the global burden of these pathologies continues to rise there is a significant need for the development of novel therapeutics. Due to their multipotent properties, stem cells have broad applications for tissue repair; additionally, stem cells have been shown to possess both immunomodulatory and neuroprotective properties. It is now believed that paracrine factors, such as extracellular vesicles (EVs), play a critical role in the functionality associated with stem cells. The diverse biological cargo contained within EVs are proposed to mediate these effects and, to date, the reparative and regenerative effects of stem cell EVs have been demonstrated in a wide range of cell types. While a high potential for their therapeutic use exists, there is a gap of knowledge surrounding their characterization, mechanisms of action, and how they may regulate cells of the CNS. Here, we report the isolation, characterization, and functional assessment of EVs from two sources of human stem cells, mesenchymal stem cells and induced pluripotent stem cells. We demonstrate the ability of these EVs to enhance the processes of cellular migration and angiogenesis, which are critical for both normal cellular development as well as cellular repair. Furthermore, we investigate their reparative effects on damaged cells, specifically those with relevance to the central nervous system. Collectively, our data highlight the similarities and differences among these EV populations and support the view that stem cells EV can be used to repair or partially reverse cellular damage. Graphical Abstract Stem cell-derived Extracellular Vesicles (EVs) for repair of damaged cells. EVs isolated from human induced pluripotent stem cells and mesenchymal stem cells contribute to the partial reversal of phenotypes induced by different sources of cellular damage.
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Affiliation(s)
- Heather Branscome
- Laboratory of Molecular Virology, School of Systems Biology, George Mason University, Discovery Hall Room 182, 10900 University Blvd, Manassas, VA, 20110, USA.,American Type Culture Collection (ATCC), Manassas, VA, USA
| | | | - Pooja Khatkar
- Laboratory of Molecular Virology, School of Systems Biology, George Mason University, Discovery Hall Room 182, 10900 University Blvd, Manassas, VA, 20110, USA
| | - Yuriy Kim
- Laboratory of Molecular Virology, School of Systems Biology, George Mason University, Discovery Hall Room 182, 10900 University Blvd, Manassas, VA, 20110, USA
| | - Robert A Barclay
- Laboratory of Molecular Virology, School of Systems Biology, George Mason University, Discovery Hall Room 182, 10900 University Blvd, Manassas, VA, 20110, USA
| | - Daniel O Pinto
- Laboratory of Molecular Virology, School of Systems Biology, George Mason University, Discovery Hall Room 182, 10900 University Blvd, Manassas, VA, 20110, USA
| | | | - Weidong Zhou
- Center for Applied Proteomics and Molecular Medicine, George Mason University, Manassas, VA, USA
| | - Lance A Liotta
- Center for Applied Proteomics and Molecular Medicine, George Mason University, Manassas, VA, USA
| | - Nazira El-Hage
- Department of Immunology, Herbert Wertheim College of Medicine, Florida International University, Miami, FL, USA
| | - Fatah Kashanchi
- Laboratory of Molecular Virology, School of Systems Biology, George Mason University, Discovery Hall Room 182, 10900 University Blvd, Manassas, VA, 20110, USA.
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14
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Intra-Vitreal Administration of Microvesicles Derived from Human Adipose-Derived Multipotent Stromal Cells Improves Retinal Functionality in Dogs with Retinal Degeneration. J Clin Med 2019; 8:jcm8040510. [PMID: 31013950 PMCID: PMC6518198 DOI: 10.3390/jcm8040510] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/14/2019] [Revised: 04/08/2019] [Accepted: 04/10/2019] [Indexed: 12/28/2022] Open
Abstract
This study was designed to determine the influence of microvesicles (MVs) derived from multipotent stromal cells isolated from human adipose tissue (hASCs) on retinal functionality in dogs with various types of retinal degeneration. The biological properties of hASC-MVs were first determined using an in vitro model of retinal Muller-like cells (CaMLCs). The in vitro assays included analysis of hASC-MVs influence on cell viability and metabolism. Brain-derived neurotrophic factor (BDNF) expression was also determined. Evaluation of the hASC-MVs was performed under normal and oxidative stress conditions. Preliminary clinical studies were performed on ten dogs with retinal degeneration. The clinical studies included behavioral tests, fundoscopy and electroretinography before and after hASC-MVs intra-vitreal injection. The in vitro study showed that CaMLCs treated with hASC-MVs were characterized by improved viability and mitochondrial potential, both under normal and oxidative stress conditions. Additionally, hASC-MVs under oxidative stress conditions reduced the number of senescence-associated markers, correlating with the increased expression of BDNF. The preliminary clinical study showed that the intra-vitreal administration of hASC-MVs significantly improved the dogs’ general behavior and tracking ability. Furthermore, fundoscopy demonstrated that the retinal blood vessels appeared to be less attenuated, and electroretinography using HMsERG demonstrated an increase in a- and b-wave amplitude after treatment. These results shed promising light on the application of cell-free therapies in veterinary medicine for retinal degenerative disorders treatment.
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15
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Liu S, Mahairaki V, Bai H, Ding Z, Li J, Witwer KW, Cheng L. Highly Purified Human Extracellular Vesicles Produced by Stem Cells Alleviate Aging Cellular Phenotypes of Senescent Human Cells. Stem Cells 2019; 37:779-790. [PMID: 30811771 PMCID: PMC6767364 DOI: 10.1002/stem.2996] [Citation(s) in RCA: 78] [Impact Index Per Article: 13.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/19/2018] [Revised: 02/07/2019] [Accepted: 02/17/2019] [Indexed: 12/21/2022]
Abstract
Extracellular vesicles (EVs), including exosomes and microvesicles, mediate intercellular communications and exert various biological activities via delivering unique cargos of functional molecules such as RNAs and proteins to recipient cells. Previous studies showed that EVs produced and secreted by human mesenchymal stem cells (MSCs) can substitute intact MSCs for tissue repair and regeneration. In this study, we examined properties and functions of EVs from human induced pluripotent stem cells (iPSCs) that can be cultured infinitely under a chemically defined medium free of any exogenous EVs. We collected and purified EVs secreted by human iPSCs and MSCs. Purified EVs produced by both stem cell types have similar sizes (∼150 nm in diameter), but human iPSCs produced 16‐fold more EVs than MSCs. When highly purified iPSC‐EVs were applied in culture to senescent MSCs that have elevated reactive oxygen species (ROS), human iPSC‐EVs reduced cellular ROS levels and alleviated aging phenotypes of senescent MSCs. Our discovery reveals that EVs from human stem cells can alleviate cellular aging in culture, at least in part by delivering intracellular peroxiredoxin antioxidant enzymes. stem cells2019;37:779–790
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Affiliation(s)
- Senquan Liu
- Department of MedicineJohns Hopkins University School of MedicineBaltimoreMarylandUSA
- Institute for Cell EngineeringJohns Hopkins University School of MedicineBaltimoreMarylandUSA
| | - Vasiliki Mahairaki
- Institute for Cell EngineeringJohns Hopkins University School of MedicineBaltimoreMarylandUSA
- Department of NeurologyJohns Hopkins University School of MedicineBaltimoreMarylandUSA
| | - Hao Bai
- Department of MedicineJohns Hopkins University School of MedicineBaltimoreMarylandUSA
- Institute for Cell EngineeringJohns Hopkins University School of MedicineBaltimoreMarylandUSA
| | - Zheng Ding
- Department of MedicineJohns Hopkins University School of MedicineBaltimoreMarylandUSA
- Institute for Cell EngineeringJohns Hopkins University School of MedicineBaltimoreMarylandUSA
| | - Jiaxin Li
- Institute for Cell EngineeringJohns Hopkins University School of MedicineBaltimoreMarylandUSA
- Department of NeurologyJohns Hopkins University School of MedicineBaltimoreMarylandUSA
| | - Kenneth W. Witwer
- Department of NeurologyJohns Hopkins University School of MedicineBaltimoreMarylandUSA
- Department of Molecular and Comparative PathobiologyJohns Hopkins University School of MedicineBaltimoreMarylandUSA
| | - Linzhao Cheng
- Department of MedicineJohns Hopkins University School of MedicineBaltimoreMarylandUSA
- Institute for Cell EngineeringJohns Hopkins University School of MedicineBaltimoreMarylandUSA
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